Figure 5.

Death induced by CTLs from granzyme AB^−/− mice. CTLs isolated from granzyme AB^−/− mice were incubated with MS9II cells. (A) Images of morphology, annexin V binding, and PI uptake were taken sequentially every 6 min. Frame a (24 min) shows the unconjugated target cell (indicated by an asterisk). By 1 h 42 min (b), the CTL (indicated by #) formed a synapse with the target (indicated by asterisks). By 2 h 12 min, (c) the target cell was blebbing and rounding, and, by 2 h 24 min (d), the target cell is fully round and still blebbing but remains annexin V and PI negative until 2 h 42 min (e). By 2 h 48 min (f), annexin V staining and PI uptake is evident. The cell continues swelling until 3 h (g). Arrow indicates the swollen plasma membrane. A montage of morphology images for each frame is presented in Fig. S1, and a full time-lapse video is presented in Video 4 (available at http://www.jcb.org/cgi/content/full/jcb.200510072/DC1). DIC, differential interference contrast. (B) Fluorescence of annexin V binding and PI relative to maximum fluorescence is presented for the cell depicted by the asterisk in A. The point at which frames (a–g) were recorded in A are shown on the time line by arrows. (C) The duration from rounding to annexin V binding and annexin V binding to PI uptake was calculated for individual cells and is presented as means ± SEM (error bars; n = 19).